There are two aspects to this work. The first is the determination of the energies required for the formation of non-covalent bonds between molecules and ions in solution. The second is to understand the relationship between the energy applied in forming a peptide ion and the type and extent of its fragmentation. Knowledge of the energetics of non-covalent bonds, particularly those involving water and biologically significant molecules, is fundamental to understanding molecular interactions and the changes in conformations that are integral to them. This research involves determining the thermodynamic quantities deltaH(std), deltaS(std), and deltaG(std)298 using the approach of equilibrium ion-molecule reaction chemistry. Hydration thermodynamics values are calculated from equilibrium constants measured over a temperature range of 0-136 degrees C at water partial pressures in the ion source ranging between zero and 100 mtorr. Equilibrium ion intensity measurements were made for at least 4 hydration states, i.e., zero through 3 water molecules associated with a core ion, and include at least 60 combinations of water partial pressures and temperatures covering the range of experimental variables. Results have been obtained for three different classes of molecules, most recently focusing on amino acids. In the past year we have completed measurements on all twenty of the essential amino acids for clusters containing up to three water molecules each. The most striking result of these extensive measurements is that, unlike other molecular or atomic ions studied previously, the enthalpy changes associated with the successive addition of waters to any of the amino acids were found to be virtually equivalent. The consequence of this is that the entropy changes for the successive waters are, unlike all previous work, dependent on the hydration state of the amino acid ion. We interpret these results to mean that amino acid hydrations are energetically very similar, but that the sites at which the water molecules are located are strongly directed by the zwitterionic nature of the ions. We have also found that the free energy for the formation of the first hydrated species is correlated significantly with the published values for amino acid hydrophobicity. In a second area of ion energetics, we have begun investigating the relationship between laser fluence and peptide ion fragmentation. This type of study is fundamental to optimizing MALDI TOF/TOF experiments for the purpose of peptide sequencing. In these studies we obtain peptide fragmentation spectra, typically 5000 laser shots, in both the unimolecular decomposition and collision induced dissociation (CID) modes. We have the ability to easily follow two time points for each peptide considered, i.e., the in-source fragmentation consisting of ions formed within 1 usec after the laser firing and the longer, mass dependent fragmentation occurring within the instrument's collision cell. To date we have studied the fragmentation of two model peptides, leucine enkephalin, YGGFL, (LeuEnk) and des-Arg-1 bradykinin, PPGFSPFR, (desRB) over the full range of laser fluence available. We observe that LeuEnk exhibits an onset of fragmentation at a much lower fluence than desRB. More importantly, however, we have been able to demonstrate that fragment formation as a function of increasing laser fluence proceeds as a set of consecutive rather than competing reactions. The implications of this are very significant for peptide sequencing since, in order to obtain a full set of fragment ions from a series of consecutive reactions, it is necessary that the peptide ion of interest be formed with sufficient internal energy to support the entire chain of decompositions. In the context of a peptide mixture, i.e., the real situation when sequencing a peptide, the necessary amount of internal energy can be supplied only in a MADI experiment. We are simultaneously developing a kinetic model for these decompositions using the Rice-Ramsberger-Kassel-Marcus (RRKM) formalism for gas phase kinetics. In addition to modeling fragmentation, the calculations will define a lower limit of the peptide ion temperatures. With this information, we will be able for the first time to estimate the fraction of laser energy delivered to gas phase ions. Furthermore, we will have a means, other than pure empiricism, to select and optimize both MALDI matrix and laser frequency.